DESCRIPTION: This project proposes the study of the structure, detailed function, and significance of the enzyme folylpolyglutamate synthetase to the chemotherapy of human tumors with folate antimetabolites. The research to be pursued entails a combination of biochemical, molecular and structural studies aimed at understanding how the enzyme converts folylmonoglutamates successively to oligo and polyglutamates. The shape of the active site will be examined and the distribution of hydrophobic and charged residues within the active site. The bioorganic chemistry of the reaction will be investigated using recombinant human enzyme and a series of probes of the active site. The structural differences apparent between the enzyme expressed in some tissues will be determined as will the molecular mechanisms responsible for control of expression of the mRNA for these species. A convergent series of techniques will be applied from expression of recombinant enzymes and active site biochemical studies to transcriptional analysis to structure-based drug design.